1. Field of the Invention
The field of this invention is lipoxygenase inhibitors.
2. Background of the Invention
Lipoxygenase are a structurally related family of nonheme iron deoxygenases that function in the production of fatty acid hydroperoxides. The lipoxygenases are present in a wide variety of organisms and catalyze the oxidation of unsaturated fatty acids utilizing an essential, non-heme iron atom. The generally accepted mechanism for lipoxygenases involves a hydrogen atom abstraction at C-3 of the 1,4-diene by Fe(III), with subsequent trapping of the pentadienyl radical by oxygen, forming the hydroperoxide product. In humans, the 5S-, 12S- and 15S-lipoxygenases oxygenate arachidonic acid in different positions along the carbon chain, e.g., as seen in leukotriene synthesis.
Lipoxygenase activity plays a critical role in asthma, atherosclerosis and cancer regulation, among other physiological processes and conditions. As such, the inhibition of lipoxygenase (LO) is currently an important goal of bio-medical research. To date, a number of different types of lipoxygenase inhibitors have been identified.
However, there is continued interest in the identification of additional lipoxygenase inhibitors.
Relevant Literature
U.S. Patents of interest include: U.S. Pat. Nos. 4,645,627 and 5,037,992. Also of interest are: Lewis et al., J. Am. Chem. Soc. (1999) 121: 1395-1396; Sailer et al., Eur. J. Biochem. (1998) 256: 364-368; and Wang et al., Biochemistry (1993) 32: 1500-1509.
Allosteric inhibitors of lipoxygenase and methods for their use are provided. In many embodiments, the allosteric inhibitors are sulfated long chain alkenyl compounds. The compounds of these embodiments are long chain monounsatured alkenyl compounds having a single sulfate moiety at the 1 position. In many embodiments, the subject compounds range in length from about 14 to 22 carbon atoms and the site of unsaturation is located between the 8 and 14 positions. Specific compounds of interest include 9-oleyl sulfate; 9-palmitoleyl sulfate and 11-eicosenyl sulfate. The subject compounds exhibit lipoxygenase inhibitory activity. As such, the subject compounds find use in treating disease conditions characterized by the presence of undesirable lipoxygenase activity. Also provided are pharmaceutical preparations of the subject compounds.
The term xe2x80x9clipoxygenasexe2x80x9d is used to refer to a protein that catalyzes the oxidation of unsaturated fatty acids utilizing an essential, non-heme iron atom. Specific lipoxygenases of interest include, but are not limited to: soybean lipoxygenase-1; 15-human lipoxygenase; 12-human lipoxygenase; 5-human lipoxygenase; 9-human lipoxygenase; and the like.
The term xe2x80x9callosteric inhibitorxe2x80x9d refers to a compound that inhibits lipoxygenase activity through binding to the allosteric binding site of the lipoxygenase protein.
xe2x80x9cAlkenylxe2x80x9d means a linear monovalent hydrocarbon chain containing at least one double bond.
xe2x80x9cLong chainxe2x80x9d means a chain having a length of from about 14 to 22 carbon atoms in length.
xe2x80x9cMonounsaturatedxe2x80x9d means a single site of unsaturation, e.g., a single double-bond between two carbon atoms in a compound.
Allosteric lipoxygenase inhibitors and methods for their use in lipoxygenase inhibition are provided. In many embodiments of the subject invention, the inhibitors are sulfated long chain alkenyl compounds. The compounds of this embodiment are generally long chain monounsatured alkenyl compounds having a single sulfate moiety at the 1 position. In many embodiments, the subject compounds range in length from about 14 to 22 carbon atoms and the site of unsaturation is located between the 8 and 14 positions. Specific compounds of interest include 9-oleyl sulfate; 9-palmitoleyl sulfate and 11-eicosenyl sulfate. The subject compounds exhibit lipoxygenase inhibitory activity. As such, the subject compounds find use in treating disease conditions characterized by the presence of undesirable lipoxygenase activity. Also provided are pharmaceutical preparations of the subject compounds. In further describing the subject invention, the subject compounds will be described first in greater detail, followed by a review of methods for their use in the lipoxygenase inhibition.
Before the subject invention is further described, it is to be understood that the invention is not limited to the particular embodiments of the invention described below, as variations of the particular embodiments may be made and still fall within the scope of the appended claims. It is also to be understood that the terminology employed is for the purpose of describing particular embodiments, and is not intended to be limiting. Instead, the scope of the present invention will be established by the appended claims.
In this specification and the appended claims, the singular forms xe2x80x9ca,xe2x80x9d xe2x80x9can,xe2x80x9d and xe2x80x9cthexe2x80x9d include plural reference unless the context clearly dictates otherwise. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this invention belongs.
Allosteric Lipoxygenase Inhibitors
As summarized above, the subject methods employ allosteric inhibitors of lipoxygenase. By allosteric inhibitor of lipoxygenase is meant a compound that binds to the allosteric binding site of lipoxygenase and at least slows, if not substantially stops, the lipoxygenase activity. As the subject compounds exhibit lipoxygenase inhibitory activity, in many embodiments the compounds exhibit a K1 value of from about 0.05 to 1.5 xcexcm, and usually from about 0.10 to 1.0 xcexcm.
Allosteric lipoxygenase inhibitors employed in the subject invention are, in many embodiments, small molecule compounds. As the inhibitors of these embodiments are small molecules, they have a molecular weight that generally ranges from about 200 to 600, usually from about 250 to 500 and more usually from about 250 to 400 daltons.
In many embodiments of the subject invention, the allosteric lipoxygenase inhibitors employed in the subject methods are sulfated long chain alkenyl compounds. By sulfated long chain alkenyl compound is meant a long chain molecule of from about 14 to 22 carbon atoms in length, where in many embodiments the number of carbon atoms in the long chain of the compounds is between 16 and 20, e.g., 16, 18 or 20. The subject compounds of this embodiment exhibit good water solubility, where their solubility typically ranges from about 150 to 200 xcexcm, usually from about 125 to 150 xcexcm and more usually from about 75 to 100 xcexcm. In addition, the subject compounds exhibit increased potency as compared to their carboxylic analogs (i.e., molecule which is the same as the subject molecule except that the sulfate moiety is replaced with a caboxylate moiety), where the magnitude of this increased potency typically ranges from about 30 to 50 fold.
As the subject compounds are alkenyl compounds, they include at least one site of unsaturation. In many embodiments, the at least one site of unsaturation is in the form of a carbon-carbon double bond. While there may be more than one site of unsaturation, in many embodiments, the compounds are monounsaturated alkenyl compounds, having a single site of unsaturation, e.g., a single carbon-carbon double bond. The site of monounsaturation typically lies between atoms 6 and 14, usually between atoms 7 and 13 and more usually between atoms 8 and 12 in the long chain alkenyl moieties of the compounds. Preferred sites of monounsaturation are positions 9 and 11 in many embodiments.
As summarized above, the subject compounds are sulfated compounds, by which is meant that they exhibit a sulfate moiety. The term xe2x80x9csulfate moietyxe2x80x9d is used to refer not only to xe2x80x94SO3H but also ionized and salt forms thereof, e.g., xe2x80x94SO3xe2x80x94; xe2x80x94SO3Na; etc. In general, the subject compounds include a single sulfate moiety located at the terminal 1 position of the compounds.
In certain embodiments of the subject invention, the employed compound are described the formula:
sulfate-(CH2)mxe2x80x94CHxe2x95x90CHxe2x80x94(CH2)nxe2x80x94CH3
wherein:
m is an integer from 7 to 10; and
n is an integer from 4 to 8.
Specific sulfated long chain alkenyl compounds of interest for use in the subject invention include, but are not limited to: 9-oleyl sulfate; 9-palmitoleyl sulfate; 11-eicosenyl sulfate; and the like.
The sulfated long chain alkenyl compounds employed in many embodiments of the subject invention may be fabricated using any convenient protocol. One representative protocol of interest is the protocol described in the Experimental Section, infra, in which a sulfate is added onto a fatty acid alcohol via a sulfamic acid/pyridine mixture. Another protocol that may be used to synthesize the subject sulfated long chain alkenyl compounds is disclosed in U.S. Pat. No. 5,037,992; the disclosure of which is herein incorporated by reference.
Methods of Lipoxygenase Inhibition
As summarized above, the subject invention provides methods of inhibiting lipoxygenase activity. By inhibiting lipoxygenase activity is meant at least a slowing, if not a substantial removal of, a protein""s lipoxygenase activity, i.e., a protein""s ability to catalyze the oxidation of unsaturated fatty acids. Where the methods result in a slowing the of the lipoxygenase activity, the degree of slowing is at least about 40 fold, usually at least about 45 fold.
In practicing the subject methods, a target lipoxygenase is contacted with an allosteric lipoxygenase inhibitor under conditions sufficient for lipoxygenase inhibition to occur. In the broadest sense, any compound that binds to the lipoxygenase allosteric binding site and consequently inhibits lipoxygenase activity may be employed. In many embodiments, the compound that is employed is a sulfated long chain alkenyl compound, as described above.
The mode of contact will vary depending on the environment of the target lipoxygenase. For example, where the target lipoxygenase is present in an in vitro environment, e.g., in a test tube, in a cell culture, etc., contact generally involves placing the allosteric inhibitor in the environment of the target lipoxygenase. Alternatively, where the target lipoxygenase is present in an in vivo environment, contact is generally achieved by administering an effective amount of the allosteric inhibitor to the host in which the in vivo environment is present. Administration may vary depending on the particular in vivo location, and may be systemic or local. A variety of different formulations and corresponding routes of administration are reviewed infra. By effective amount is meant an amount effective to cause the desired amount of lipoxygenase inhibition. The amount administered to a host may vary significantly depending on the nature of the compound, the nature of the host, the location of the target lipoxygenase, the route of administration, etc., where the dosages may readily be determined empirically by those of skill in the art.
The subject methods of inhibiting lipoxygenase activity find use in a variety of different applications, including the treatment of a variety of different disease conditions associated with lipoxygenase activity in a variety of different hosts. Representative conditions in which the subject methods find use include, but are not limited to: the treatment of asthma, atherosclerosis, cellular proliferative diseases, e.g., cancer, psoriasis, etc., inflammation, bone resorption suppression, removal of hair (See U.S. Pat. No. 5,928,654, dislosure of which is herein incorporated by reference), and the like.
By treatment is meant at least an amelioration in the symptoms of the disease conditions experienced by the host, where treatment can include substantially complete cessation, if not complete cessation of symptoms and/or substantially complete removal, if not complete removal of the underlying cause of the symptoms.
As mentioned above, the compounds of the subject invention find use in treating conditions in a variety of different hosts. Generally such hosts are xe2x80x9cmammalsxe2x80x9d or xe2x80x9cmammalian,xe2x80x9d where these terms are used broadly to describe organisms which are within the class mammalia, including the orders carnivore (e.g., dogs and cats), rodentia (e.g., mice, guinea pigs, and rats), and primates (e.g., humans, chimpanzees, and monkeys). In many embodiments, the hosts will be humans.
Pharmaceutical Preparations
Also provided are pharmaceutical preparations of the subject allosteric lipoxygenase inhibitor compounds. The subject compounds can be incorporated into a variety of formulations for therapeutic administration. More particularly, the compounds of the present invention can be formulated into pharmaceutical compositions by combination with appropriate, pharmaceutically acceptable carriers or diluents, and may be formulated into preparations in solid, semi-solid, liquid or gaseous forms, such as tablets, capsules, powders, granules, ointments, solutions, suppositories, injections, inhalants and aerosols. The formulations may be designed for administration via a number of different routes, including oral, buccal, rectal, parenteral, intraperitoneal, intradermal, transdermal, intracheal, etc., administration.
In pharmaceutical dosage forms, the compounds may be administered in the form of their pharmaceutically acceptable salts, or they may also be used alone or in appropriate association, as well as in combination, with other pharmaceutically active compounds. The following methods and excipients are merely exemplary and are in no way limiting.
For oral preparations, the compounds can be used alone or in combination with appropriate additives to make tablets, powders, granules or capsules, for example, with conventional additives, such as lactose, mannitol, corn starch or potato starch; with binders, such as crystalline cellulose, cellulose derivatives, acacia, corn starch or gelatins; with disintegrators, such as corn starch, potato starch or sodium carboxymethylcellulose; with lubricants, such as talc or magnesium stearate; and if desired, with diluents, buffering agents, moistening agents, preservatives and flavoring agents.
The compounds can be formulated into preparations for injection by dissolving, suspending or emulsifying them in an aqueous or nonaqueous solvent, such as vegetable or other similar oils, synthetic aliphatic acid glycerides, esters of higher aliphatic acids or propylene glycol; and if desired, with conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifying agents, stabilizers and preservatives.
The compounds can be utilized in aerosol formulation to be administered via inhalation. The compounds of the present invention can be formulated into pressurized acceptable propellants such as dichlorodifluoromethane, propane, nitrogen and the like.
Furthermore, the compounds can be made into suppositories by mixing with a variety of bases such as emulsifying bases or water-soluble bases. The compounds of the present invention can be administered rectally via a suppository. The suppository can include vehicles such as cocoa butter, carbowaxes and polyethylene glycols, which melt at body temperature, yet are solidified at room temperature.
Unit dosage forms for oral or rectal administration such as syrups, elixirs, and suspensions may be provided wherein each dosage unit, for example, teaspoonful, tablespoonful, tablet or suppository, contains a predetermined amount of the composition containing one or more inhibitors. Similarly, unit dosage forms for injection or intravenous administration may comprise the inhibitor(s) in a composition as a solution in sterile water, normal saline or another pharmaceutically acceptable carrier.
The term xe2x80x9cunit dosage form,xe2x80x9d as used herein, refers to physically discrete units suitable as unitary dosages for human and animal subjects, each unit containing a predetermined quantity of compounds of the present invention calculated in an amount sufficient to produce the desired effect in association with a pharmaceutically acceptable diluent, carrier or vehicle. The specifications for the novel unit dosage forms of the present invention depend on the particular compound employed and the effect to be achieved, and the pharmacodynamics associated with each compound in the host.
The pharmaceutically acceptable excipients, such as vehicles, adjuvants, carriers or diluents, are readily available to the public. Moreover, pharmaceutically acceptable auxiliary substances, such as pH adjusting and buffering agents, tonicity adjusting agents, stabilizers, wetting agents and the like, are readily available to the public.
Those of skill will readily appreciate that dose levels can vary as a function of the specific compound, the severity of the symptoms and the susceptibility of the subject to side effects. Preferred dosages for a given compound are readily determinable by those of skill in the art by a variety of means.
Kits with unit doses of the compounds, usually in oral, injectable or topical doses, are provided. In such kits, in addition to the containers containing the unit doses will be an informational package insert describing the use and attendant benefits of the drugs in treating pathological condition of interest. Preferred compounds and unit doses are those described herein above.